GENETIC DIFFERENTIATION AND SELECTION AGAINST MIGRANTS IN EVOLUTIONARILY REPLICATED EXTREME ENVIRONMENTS

We investigated mechanisms of reproductive isolation in livebearing fishes (genus Poecilia) inhabiting sulfidic and nonsulfidic habitats in three replicate river drainages. Although sulfide spring fish convergently evolved divergent phenotypes, it was unclear if mechanisms of reproductive isolation also evolved convergently. Using microsatellites, we found strongly reduced gene flow between adjacent populations from different habitat types, suggesting that local adaptation to sulfidic habitats repeatedly caused the emergence of reproductive isolation. Reciprocal translocation experiments indicate strong selection against immigrants into sulfidic waters, but also variation among drainages in the strength of selection against immigrants into nonsulfidic waters. Mate choice experiments revealed the evolution of assortative mating preferences in females from nonsulfidic but not from sulfidic habitats. The inferred strength of sexual selection against immigrants (RI_s) was negatively correlated with the strength of natural selection (RI_m), a pattern that could be attributed to reinforcement, whereby natural selection strengthens behavioral isolation due to reduced hybrid fitness. Overall, reproductive isolation and genetic differentiation appear to be replicated and direct consequences of local adaptation to sulfide spring environments, but the relative contributions of different mechanisms of reproductive isolation vary across these evolutionarily independent replicates, highlighting both convergent and nonconvergent evolutionary trajectories of populations in each drainage.     

Divergence in trophic ecology characterizes colonization of extreme habitats

Extreme habitats are characterized by the presence of physio‐chemical stressors, but also differ in aspects of the biotic environment, such as resource availability or the presence of competitors. The present study quantifies variation in trophic ecology of a small livebearing fish (Poecilia mexicana, Poeciliidae) across four different habitats that included nonsulphidic and sulphidic surface waters, as well as a nonsulphidic and a sulphidic cave. Resource use in different habitat types was investigated using gut content analysis. Populations diverged in resource use from a diet dominated by algae and detritus in nonsulfidic surface habitats to a diet including invertebrate food items in the other habitats. Poecilia mexicana in cave habitats further exhibited a higher dietary niche width than conspecifics from surface habitats. The condition of P. mexicana was analysed using storage lipid extractions. Fish from sulphidic and cave habitats exhibited a very poor condition, suggesting resource limitation and/or high costs of coping with extreme conditions. Finally, divergence in resource use was correlated with variation in viscerocranial morphology. A common garden experiment indicated both a genetic and plastic basis to the morphological variation observed among field populations. It is suggested that the morphological diversification is an adaptation to the differential use of resources among populations. © 2008 The Linnean Society of London, Biological Journal of the Linnean Society, 2008, 95 , 517–528.     

Extreme habitats are not refuges: poeciliids suffer from increased aerial predation risk in sulphidic southern Mexican habitats

Extreme environments are often considered a predation refuge for organisms living in them. In southern Mexico several species of poeciliid fishes are undergoing incipient speciation in a variety of extreme (i.e. permanently dark and/or sulphidic) freshwater systems, and previous research has demonstrated reproductive isolation between populations from sulphidic and adjacent benign habitats. In the present study, we investigated bird predation rates (measured as successful captures per minute) in two sulphidic surface and several benign surface habitats, to test the hypothesis that extreme habitats are predation refuges. We found capture rates to be approximately 20 times higher in sulphidic environments: probably facilitated by extremophile poeciliids spending most of their time at the water surface, where they engage in aquatic surface respiration as a direct response to hypoxia. Even birds that are usually not considered major fish predators regularly engage in fish predation in the toxic habitats of southern Mexico. Our results demonstrate that extreme environments do not necessarily represent a refuge from predation, and we discuss the general importance of predation in driving incipient speciation in these systems. Finally, we hypothesize that natural selection via avian predation may play an important role in maintaining reproductive isolation between divergent poeciliid populations. © 2010 The Linnean Society of London, Biological Journal of the Linnean Society, 2010, 101 , 417–426.     

Perspective: Reproductive isolation caused by natural selection against immigrants from divergent habitats

The classification of reproductive isolating barriers laid out by Dobzhansky and Mayr has motivated and structured decades of research on speciation. We argue, however, that this classification is incomplete and that the unique contributions of a major source of reproductive isolation have often been overlooked. Here, we describe reproductive barriers that derive from the reduced survival of immigrants upon reaching foreign habitats that are ecologically divergent from their native habitat. This selection against immigrants reduces encounters and thus mating opportunities between individuals from divergently adapted populations. It also reduces the likelihood that successfully mated immigrant females will survive long enough to produce their hybrid offspring. Thus, natural selection against immigrants results in distinctive elements of premating and postmating reproductive isolation that we hereby dub "immigrant inviability." We quantify the contributions of immigrant inviability to total reproductive isolation by examining study systems where multiple components of reproductive isolation have been measured and demonstrate that these contributions are frequently greater than those of traditionally recognized reproductive barriers. The relevance of immigrant inviability is further illustrated by a consideration of population-genetic theory, a review of selection against immigrant alleles in hybrid zone studies, and an examination of its participation in feedback loops that influence the evolution of additional reproductive barriers. Because some degree of immigrant inviability will commonly exist between populations that exhibit adaptive ecological divergence, we emphasize that these barriers play critical roles in ecological modes of speciation. We hope that the formal recognition of immigrant inviability and our demonstration of its evolutionary importance will stimulate more explicit empirical studies of its contributions to speciation.     

The evolution of host preference in allopatric vs. parapatric populations of Timema cristinae walking-sticks

Divergent habitat preferences can contribute to speciation, as has been observed for host-plant preferences in phytophagous insects. Geographic variation in host preference can provide insight into the causes of preference evolution. For example, selection against maladaptive host-switching occurs only when multiple hosts are available in the local environment and can result in greater divergence in regions with multiple vs. a single host. Conversely, costs of finding a suitable host can select for preference even in populations using a single host. Some populations of Timema cristinae occur in regions with only one host-plant species present (in allopatry, surrounded by unsuitable hosts) whereas others occur in regions with two host-plant species adjacent to one another (in parapatry). Here, we use host choice and reciprocal-rearing experiments to document genetic divergence in host preference among 33 populations of T. cristinae. Populations feeding on Ceanothus exhibited a stronger preference for Ceanothus than did populations feeding on Adenostoma. Both allopatric and parapatric pairs of populations using the different hosts exhibited divergent host preferences, but the degree of divergence tended to be greater between allopatric pairs. Thus, gene flow between parapatric populations apparently constrains divergence. Host preferences led to levels of premating isolation between populations using alternate hosts that were comparable in magnitude to previously documented premating isolation caused by natural and sexual selection against migrants between hosts. Our findings demonstrate how gene flow and different forms of selection interact to determine the magnitude of reproductive isolation observed in nature.     

Adaptation with gene flow across the landscape in a dune sunflower

Isolation by adaptation increases divergence at neutral loci when natural selection against immigrants reduces the rate of gene flow between different habitats. This can occur early in the process of adaptive divergence and is a key feature of ecological speciation. Despite the ability of isolation by distance (IBD) and other forms of landscape resistance to produce similar patterns of neutral divergence within species, few studies have used landscape genetics to control for these other forces. We have studied the divergence of Helianthus petiolaris ecotypes living in active sand dunes and adjacent non-dune habitat, using landscape genetics approaches, such as circuit theory and multiple regression of distance matrices, in addition to coalescent modelling. Divergence between habitats was significant, but not strong, and was shaped by IBD. We expected that increased resistance owing to patchy and unfavourable habitat in the dunes would contribute to divergence. Instead, we found that landscape resistance models with lower resistance in the dunes performed well as predictors of genetic distances among subpopulations. Nevertheless, habitat class remained a strong predictor of genetic distance when controlling for isolation by resistance and IBD. We also measured environmental variables at each site and confirmed that specific variables, especially soil nitrogen and vegetation cover, explained a greater proportion of variance in genetic distance than did landscape or the habitat classification alone. Asymmetry in effective population sizes and numbers of migrants per generation was detected using coalescent modelling with Bayesian inference, which is consistent with incipient ecological speciation being driven by the dune habitat.     

Toxic hydrogen sulfide and dark caves: phenotypic and genetic divergence across two abiotic environmental gradients in Poecilia mexicana

Divergent natural selection drives evolutionary diversification. It creates phenotypic diversity by favoring developmental plasticity within populations or genetic differentiation and local adaptation among populations. We investigated phenotypic and genetic divergence in the livebearing fish Poecilia mexicana along two abiotic environmental gradients. These fish typically inhabit nonsulfidic surface rivers, but also colonized sulfidic and cave habitats. We assessed phenotypic variation among a factorial combination of habitat types using geometric and traditional morphometrics, and genetic divergence using quantitative and molecular genetic analyses. Fish in caves (sulfidic or not) exhibited reduced eyes and slender bodies. Fish from sulfidic habitats (surface or cave) exhibited larger heads and longer gill filaments. Common-garden rearing suggested that these morphological differences are partly heritable. Population genetic analyses using microsatellites as well as cytochrome b gene sequences indicate high population differentiation over small spatial scale and very low rates of gene flow, especially among different habitat types. This suggests that divergent environmental conditions constitute barriers to gene flow. Strong molecular divergence over short distances as well as phenotypic and quantitative genetic divergence across habitats in directions classic to fish ecomorphology suggest that divergent selection is structuring phenotypic variation in this system.     

The relative influence of natural selection and geography on gene flow in guppies

Two general processes may influence gene flow among populations. One involves divergent selection, wherein the maladaptation of immigrants and hybrids impedes gene flow between ecological environments (i.e. ecological speciation). The other involves geographic features that limit dispersal. We determined the relative influence of these two processes in natural populations of Trinidadian guppies (Poecilia reticulata). If selection is important, gene flow should be reduced between different selective environments. If geography is important, gene flow should be impeded by geographic distance and physical barriers. We examined how genetic divergence, long-term gene flow, and contemporary dispersal within a watershed were influenced by waterfalls, geographic distance, predation, and habitat features. We found that waterfalls and geographic distance increased genetic divergence and reduced dispersal and long-term gene flow. Differences in predation or habitat features did not influence genetic divergence or gene flow. In contrast, differences in predation did appear to reduce contemporary dispersal. We suggest that the standard predictions of ecological speciation may be heavily nuanced by the mating behaviour and life history strategies of guppies.     

Batesian mimicry promotes pre‐ and postmating isolation in a snake mimicry complex

We evaluated whether Batesian mimicry promotes early‐stage reproductive isolation. Many Batesian mimics occur not only in sympatry with their model (as expected), but also in allopatry. As a consequence of local adaptation within both sympatry (where mimetic traits are favored) and allopatry (where nonmimetic traits are favored), divergent, predator‐mediated natural selection should disfavor immigrants between these selective environments as well as any between‐environment hybrids. This selection might form the basis for both pre‐ and postmating isolation, respectively. We tested for such selection in a snake mimicry complex by placing clay replicas of sympatric, allopatric, or hybrid phenotypes in both sympatry and allopatry and measuring predation attempts. As predicted, replicas with immigrant phenotypes were disfavored in both selective environments. Replicas with hybrid phenotypes were also disfavored, but only in a region of sympatry where previous studies have detected strong selection favoring precise mimicry. By fostering immigrant inviability and ecologically dependent selection against hybrids (at least in some habitats), Batesian mimicry might therefore promote reproductive isolation. Thus, although Batesian mimicry has long been viewed as a mechanism for convergent evolution, it might play an underappreciated role in fueling divergent evolution and possibly even the evolution of reproductive isolation and speciation.     

Partitioning the effects of isolation by distance,environment, and physical barriers on genomic divergence between parapatric threespine stickleback

Genetic divergence between populations is shaped by a combination of drift, migration, and selection, yielding patterns of isolation‐by‐distance (IBD) and isolation‐by‐environment (IBE). Unfortunately, IBD and IBE may be confounded when comparing divergence across habitat boundaries. For instance, parapatric lake and stream threespine stickleback (Gasterosteus aculeatus) may have diverged due to selection against migrants (IBE), or mere spatial separation (IBD). To quantitatively partition the strength of IBE and IBD, we used recently developed population genetic software (BEDASSLE) to analyze partial genomic data from three lake‐stream clines on Vancouver Island. We find support for IBD within each of three outlet streams (unlike prior studies of lake‐stream stickleback). In addition, we find evidence for IBE (controlling for geographic distance): the genetic effect of habitat is equivalent to geographic separation of ～1.9 km of IBD. Remarkably, of our three lake‐stream pairs, IBE is strongest where migration between habitats is easiest. Such microgeographic genetic divergence would require exceptionally strong divergent selection, which multiple experiments have failed to detect. Instead, we propose that nonrandom dispersal (e.g., habitat choice) contributes to IBE. Supporting this conclusion, we show that the few migrants between habitats are a nonrandom subset of the phenotype distribution of the source population.     

Recent divergence with gene flow in Tennessee cave salamanders (Plethodontidae: Gyrinophilus) inferred from gene genealogies

Cave organisms occupy a special place in evolutionary biology because convergent morphologies of many species demonstrate repeatability in evolution even as they obscure phylogenetic relationships. The origin of specialized cave-dwelling species also raises the issue of the relative importance of isolation vs. natural selection in speciation. Two alternative hypotheses describe the origin of subterranean species. The 'climate-relict' model proposes allopatric speciation after populations of cold-adapted species become stranded in caves due to climate change. The 'adaptive-shift' model proposes parapatric speciation driven by divergent selection between subterranean and surface habitats. Our study of the Tennessee cave salamander complex shows that the three nominal forms (Gyrinophilus palleucus palleucus, G. p. necturoides, and G. gulolineatus) arose recently and are genealogically nested within the epigean (surface-dwelling) species, G. porphyriticus. Short branch lengths and discordant gene trees were consistent with a complex history involving gene flow between diverging forms. Results of coalescent-based analysis of the distribution of haplotypes among groups reject the allopatric speciation model and support continuous or recurrent genetic exchange during divergence. These results strongly favour the hypothesis that Tennessee cave salamanders originated from spring salamanders via divergence with gene flow.     

Assessing reproductive isolation using a contact zone between parapatric lake‐stream stickleback ecotypes

Ecological speciation occurs when populations evolve reproductive isolation as a result of divergent natural selection. This isolation can be influenced by many potential reproductive barriers, including selection against hybrids, selection against migrants and assortative mating. How and when these barriers act and interact in nature is understood for relatively few empirical systems. We used a mark–recapture experiment in a contact zone between lake and stream three‐spined sticklebacks (Gasterosteus aculeatus, Linnaeus) to evaluate the occurrence of hybrids (allowing inferences about mating isolation), the interannual survival of hybrids (allowing inferences about selection against hybrids) and the shift in lake‐like vs. stream‐like characteristics (allowing inferences about selection against migrants). Genetic and morphological data suggest the occurrence of hybrids and no selection against hybrids in general, a result contradictory to a number of other studies of sticklebacks. However, we did find selection against more lake‐like individuals, suggesting a barrier to gene flow from the lake into the stream. Combined with previous work on this system, our results suggest that multiple (most weakly and often asymmetric) barriers must be combining to yield substantial restrictions on gene flow. This work provides evidence of a reproductive barrier in lake–stream sticklebacks and highlights the value of assessing multiple reproductive barriers in natural contexts.     

Does a predatory insect contribute to the divergence between cave- and surface-adapted fish populations?

Immigrant inviability, where individuals from foreign, ecologically divergent habitats are less likely to survive, can restrict gene flow among diverging populations and result in speciation. I investigated whether a predatory aquatic insect (Belostoma sp.) selects against migrants between cave and surface populations of a fish (Poecilia mexicana). Cavefish were more susceptible to attacks in the light, whereas surface fish were more susceptible in darkness. Environmentally dependent susceptibility to attacks may thus contribute to genetic and phenotypic differentiation between the populations. This study highlights how predation—in this case in conjunction with differences in other environmental factors—can be an important driver in speciation.     

Ecological reproductive isolation of coast and inland races of Mimulus guttatus

Adaptive divergence due to habitat differences is thought to play a major role in formation of new species. However it is rarely clear the extent to which individual reproductive isolating barriers related to habitat differentiation contribute to total isolation. Furthermore, it is often difficult to determine the specific environmental variables that drive the evolution of those ecological barriers, and the geographic scale at which habitat-mediated speciation occurs. Here, we address these questions through an analysis of the population structure and reproductive isolation between coastal perennial and inland annual forms of the yellow monkeyflower, Mimulus guttatus. We found substantial morphological and molecular genetic divergence among populations derived from coast and inland habitats. Reciprocal transplant experiments revealed nearly complete reproductive isolation between coast and inland populations mediated by selection against immigrants and flowering time differences, but not postzygotic isolation. Our results suggest that selection against immigrants is a function of adaptations to seasonal drought in inland habitat and to year round soil moisture and salt spray in coastal habitat. We conclude that the coast and inland populations collectively comprise distinct ecological races. Overall, this study suggests that adaptations to widespread habitats can lead to the formation of reproductively isolated species.     

Correlated divergence of female and male genitalia in replicated lineages with ongoing ecological speciation

Divergence of genital traits among lineages has the potential to serve as a reproductive isolating barrier when copulation, insemination, and fertilization are inhibited by incompatibilities between female and male genitalia. Despite widespread evidence for genital trait diversity among closely related lineages and coevolution of female and male genitalia within lineages, few studies have investigated genital evolution during the early stages of speciation. We quantified genital variation in replicated population pairs of Poecilia mexicana with ongoing ecological speciation between sulfidic (H₂S containing) and nearby nonsulfidic habitats. These analyses revealed rapid and correlated divergence of female and male genitalia across evolutionarily independent population pairs exposed to divergent selection regimes. Both sexes exhibited convergent evolution of genital traits among populations inhabiting similar habitat types. Our results demonstrate that genital evolution can occur during the early stages of speciation‐with‐gene‐flow, potentially as a result of variation in the intensity of sexual conflict among populations. Our results suggest genitalia may contribute to early stages of divergence and challenge the generality of previously suggested mechanisms of genital evolution in poeciliids.     

Morphological divergence within and between the Mexican sailfin mollies, Poecilia velifera and Poecilia petenensis

This study examined the patterns of morphological variation both between species, and between sexes and among populations within each species of the Mexican sailfin mollies, Poecilia velifera and Poecilia petenensis , using geometric morphometrics and linear measurements of morphological traits. While sexes within each species differed in characteristics that may be important in sexual selection, such as length of the dorsal fin, species differed in traits, such as body depth, that may also be influenced by natural selection due to differences in habitats. Within each species, many morphological traits were similar among populations, but important differences, including caudal peduncle depth in P. petenensis (but not in P. velifera ), suggested that habitat differences may also be important in shaping population divergence independently within each species. Indeed, the evolutionary vectors of male morphological population divergence for each species differed by an angle of 98·5°, representing nearly orthogonal vectors and suggesting independent shape divergence between these two molly species. Finally, geographic isolation did not explain the morphological differentiation seen among populations, suggesting that natural and sexual selection were strong forces promoting morphological diversification within these two species, despite the potential for a high degree of population connectivity and gene flow.     

The consequences of phenotypic plasticity for ecological speciation

We use an individual-based numerical simulation to study the effects of phenotypic plasticity on ecological speciation. We find that adaptive plasticity evolves readily in the presence of dispersal between populations from different ecological environments. This plasticity promotes the colonization of new environments but reduces genetic divergence between them. We also find that the evolution of plasticity can either enhance or degrade the potential for divergent selection to form reproductive barriers. Of particular importance here is the timing of plasticity in relation to the timing of dispersal. If plasticity is expressed after dispersal, reproductive barriers are generally weaker because plasticity allows migrants to be better suited for their new environment. If plasticity is expressed before dispersal, reproductive barriers are either unaffected or enhanced. Among the potential reproductive barriers we considered, natural selection against migrants was the most important, primarily because it was the earliest-acting barrier. Accordingly, plasticity had a much greater effect on natural selection against migrants than on sexual selection against migrants or on natural and sexual selection against hybrids. In general, phenotypic plasticity can strongly alter the process of ecological speciation and should be considered when studying the evolution of reproductive barriers.     

Effects of natural and sexual selection on adaptive population divergence and premating isolation in a damselfly

The relative strength of different types of directional selection has seldom been compared directly in natural populations. A recent meta-analysis of phenotypic selection studies in natural populations suggested that directional sexual selection may be stronger in magnitude than directional natural selection, although this pattern may have partly been confounded by the different time scales over which selection was estimated. Knowledge about the strength of different types of selection is of general interest for understanding how selective forces affect adaptive population divergence and how they may influence speciation. We studied divergent selection on morphology in parapatric, natural damselfly (Calopteryx splendens) populations. Sexual selection was stronger than natural selection measured on the same traits, irrespective of the time scale over which sexual selection was measured. Visualization of the fitness surfaces indicated that population divergence in overall morphology is more strongly influenced by divergent sexual selection rather than natural selection. Courtship success of experimental immigrant males was lower than that of resident males, indicating incipient sexual isolation between these populations. We conclude that current and strong sexual selection promotes adaptive population divergence in this species and that premating sexual isolation may have arisen as a correlated response to divergent sexual selection. Our results highlight the importance of sexual selection, rather than natural selection in the adaptive radiation of odonates, and supports previous suggestions that divergent sexual selection promotes speciation in this group.     

VARIABLE PROGRESS TOWARD ECOLOGICAL SPECIATION IN PARAPATRY: STICKLEBACK ACROSS EIGHT LAKE-STREAM TRANSITIONS

Divergent selection between contrasting habitats can sometimes drive adaptive divergence and the evolution of reproductive isolation in the face of initially high gene flow. "Progress" along this ecological speciation pathway can range from minimal divergence to full speciation. We examine this variation for threespine stickleback fish that evolved independently across eight lake-stream habitat transitions. By quantifying stickleback diets, we show that lake-stream transitions usually coincide with limnetic-benthic ecotones. By measuring genetically based phenotypes, we show that these ecotones often generate adaptive divergence in foraging morphology. By analyzing neutral genetic markers (microsatellites), we show that adaptive divergence is often associated with the presence of two populations maintaining at least partial reproductive isolation in parapatry. Coalescent-based simulations further suggest that these populations have diverged with gene flow within a few thousand generations, although we cannot rule out the possibility of phases of allopatric divergence. Finally, we find striking variation among the eight lake-stream transitions in progress toward ecological speciation. This variation allows us to hypothesize that progress is generally promoted by strong divergent selection and limited dispersal across the habitat transitions. Our study thus makes a case for ecological speciation in a parapatric context, while also highlighting variation in the outcome.